We continued biochemical studies of mammalian DNA replication proteins. Our full length cDNA for human beta-polymerase was subcloned in an expression vector. The protein was overproduced in E. coli and purified in mg quantities. Enzymological characterization indicated that the recombinant enzyme is similar to the natural enzyme in catalytic properties. In vitro DNA repair activities of the recombinant enzyme were studied. Genomic clones spanning the gene for human beta-polymerase were isolated and characterized. The promoter region was sequenced and studied by deletion mutagenesis using a transient system. In other work, we found that the abundance of beta- polymerase mRNA in cultured human cells in both cell cycle regulated and serum regulated. The beta-polymerase coding sequence was stably transfected into human fibroblast and expression of the transfected gene was studied. Steady-state kinetic analysis of the HIV DNA polymerase was conducted, and an overall kinetic scheme was derived. A DNA segment containing the coding region for this enzyme was subcloned in an expression vector and the enzyme was over- produced in E. coli. Physical biochemical studies of a recombinant single-stranded nucleic acid binding protein termed A1 were conducted. The protein binds cooperatively to either RNA or DNA and the full- length protein binds much tighter than a truncated Al protein lacking the glycine-rich COOH-terminal domain (residues 185- 319). Proton NMR studies suggest that mechanism of Al binding is similar to that of several prokaryotic ssDNA binding proteins in that binding involves close approach of aromatic amino acids with nucleotide bases. Indeed, we found that all 4 major sites of covalent Al photocross-linking to (32P)d(pT)8 occur at Phe residues. No crosslinking sites were found within the COOH- terminal domain, yet this domain clearly makes a significant contribution to the overall free energy of binding of Al to nucleic acids.